Matrix Metalloproteinases and their natural inhibitors (TIMPs) regulate the dissolution of the extracellular matrix in growth and development, inflammatory and neoplastic diseases. The objective of this project is to investigate how cells orchestrate the episodic local dissolution of extracellular matrix by the orderly expression and function of MMPs and their inhibitors. Initially our studies are aimed at determining which MMPs are involved in the degradation of a single substrate (reconstituted fibrils of type 1 collagen) by a single cell type. To address this question we are employing a variety of approaches including enzyme identification techniques by neo- epitope finger-printing of substrate cleavage sites, blocking by group selective synthetic inhibitors or by enzyme-specific antibodies. In order to monitor the progression of conversion of MMP zymogens (primarily MMP-1) to the catalytically active form, antibodies which can recognize and discriminate between the two forms are being developed. A body of evidence suggests that MMP activity and MMP zymogen activation are regulated and controlled by TIMPs. To assess the role and function of TIMPs (specifically TIMP-1 and TIMP- 2) we are developing strategies aimed at replacement of the murine TIMP-1 and/or TIMP-2 genes with either null mutants, conditional null mutants or mutants with altered function.